LOCUS       sequence-232330-        6159 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  shuttle vector for baculovirus production, using FlashBac bacmid; 
            for recombinant protein with N-terminal HIS6 tag, cleavable with 3C,
            and C-terminal monomeric GFP, cleavable with TEV.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6159)
  AUTHORS   Lemaitre RP, Bogdanova A, Borgonovo B, Woodruff JB, Drechsel DN
  TITLE     FlexiBAC: a versatile, open-source baculovirus vector system for 
            protein expression, secretion, and proteolytic processing.
  JOURNAL   BMC Biotechnol. 2019 Mar 29;19(1):20. doi: 
            10.1186/s12896-019-0512-z.
  PUBMED    30925874
REFERENCE   2  (bases 1 to 6159)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "BMC 
            Biotechnol."; date: "2019-03-29"; volume: "19(1):20. doi"; pages: " 
            10.1186/s12896-019-0512-z"
COMMENT     Sequence Label: pOCC69
FEATURES             Location/Qualifiers
     source          1..6159
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..92
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     17..34
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
     regulatory      134..143
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             152..169
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             182..205
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             314..619
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     498..517
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             675..695
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             join(696..698,699..701,702..1415)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(741..762)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1002..1021)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1349..1370
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1548..1682
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1585..1604)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1639..1658
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     3'UTR           1755..2127
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     primer_bind     complement(2479..2496)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2589..3177)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2669..2688)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3348..4139,4140..4208))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     3971..3990
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4209..4313)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4614..4636
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4628..4645
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4629..4645
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4656..4675
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4656..4674
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             5285..5409
                     /codon_start=3
                     /product="baculovirus late expression factor 2"
                     /label=lef2
                     /translation="HLLIKSQDVYKPPNCQKMKTVDKLCPFAGNCKGLNPICNY"
     CDS             complement(5447..6052)
                     /codon_start=1
                     /product="baculovirus ORF603 protein"
                     /label=ORF603
                     /translation="MAVIFNNKQLLADNSIEKGGELFLFNGSYNILESYVNPVLLKNGV
                     IELEEAAYYAGNILYKTDDPKFIDYINLIIKATHSEELPENSTVVNYRKTMRSGTIHPI
                     KKDIYIYDNKKFTLYDRYIYGYDNNYVNFYEEKNEKEKEYEEEDDKASSLCENKIILSQ
                     INCESFENDFKYYLSDYNYAFSIIDNTTNVLVAFGLYR"
ORIGIN
        1 atcatggaga taattaaaat gataaccatc tcgcaaataa ataagtattt tactgttttc
       61 gtaacagttt tgtaataaaa aaacctataa atattccgga ttattcatac cgtcccacca
      121 tcgggcgcgg atcgccacca tgggcagcag ccatcaccat caccatcaca gcagcggcag
      181 actggaagtt ctgttccagg ggccggcggc cgcaagcgct atggccacaa cctgttacat
      241 tgcacaagat aaaaatatat catcacgaac agtaaaactg tctgcttaca taaacagtaa
      301 tacaaggggt gttatgcagt ttaaggttta cacctataaa agagagagcc gttatcgtct
      361 gtttgtggat gtacagagtg atattattga cacgcccggg cgacggatgg tgatccccct
      421 ggccagtgcc cgtctgctgt cagataaagt ctcccgtgaa ctttacccgg tggtgcatat
      481 cggggatgaa agctggcgca tgatgaccac cgatatggcc agtgtgccgg tctccgttat
      541 cggggaagaa gtggctgatc tcagccaccg cgaaaatgac atcaaaaacg ccattaacct
      601 gatgttctgg ggaatataaa tgtcaggctc ccttatacac agccagtggc gcgccgggat
      661 ccagcagcgg cagagaaaac ttgtatttcc agggcatggt gagcaagggc gaggagctgt
      721 tcaccggggt ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc cacaagttca
      781 gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa gctgaccctg aagttcatct
      841 gcaccaccgg caagctgccc gtgccctggc ccaccctcgt gaccaccctg acctacggcg
      901 tgcagtgctt cagccgctac cccgaccaca tgaagcagca cgacttcttc aagtccgcca
      961 tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa ggacgacggc aactacaaga
     1021 cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca
     1081 tcgacttcaa ggaggacggc aacatcctgg ggcacaagct ggagtacaac tacaacagcc
     1141 acaacgtcta tatcatggcc gacaagcaga agaacggcat caaggtgaac ttcaagatcc
     1201 gccacaacat cgaggacggc agcgtgcagc tcgccgacca ctaccagcag aacaccccca
     1261 tcggcgacgg ccccgtgctg ctgcccgaca accactacct gagcacccag tccaaactga
     1321 gcaaagaccc caacgagaag cgcgatcaca tggtcctgct ggagttcgtg accgccgccg
     1381 ggatcactct cggcatggac gagctgtaca agtaatagaa gcttgtcgag aagtactaga
     1441 ggatcataat cagccatacc acatttgtag aggttttact tgctttaaaa aacctcccac
     1501 acctccccct gaacctgaaa cataaaatga atgcaattgt tgttgttaac ttgtttattg
     1561 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt
     1621 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgga
     1681 tctgatcact gcttgagcct aggagatccg aaccagataa gtgaaagcct gcatccgcag
     1741 atcttaatta ataaaacacg atacattgtt attagtacat ttattaagcg ctagattctg
     1801 tgcgttgttg atttacagac aattgttgta cgtattttaa taattcatta aatttataat
     1861 ctttagggtg gtatgttaga gcgaaaatca aatgattttc agcgtcttta tatctgaatt
     1921 taaatattaa atcctcaata gatttgtaaa ataggtttcg attagtttca aacaagggtt
     1981 gtttttccga accgatggct ggactatcta atggattttc gctcaacgcc acaaaacttg
     2041 ccaaatcttg tagcagcaat ctagctttgt cgatattcgt ttgtgttttg ttttgtaata
     2101 aaggttcgac gtcgttcaaa atattatgcg cttttgtatt tctttcatca ctgtcgttag
     2161 tgtacaattg actcgacgta aacacgttaa ataacgcttg gacatattta acatcgggcg
     2221 tgttagcttt attaggccga ttatcgtcgt cgtcccaacc ctcgtcgtta gaagttgctt
     2281 ccgaagacga ttttgccata gccacacgac gcctattaat tgtgtcggct aacacgtccg
     2341 cgatcaaatt tgtagttgag ctttttgggc ctcatgggcc ttccgctcac tgcccgcttt
     2401 ccagtcggga aacctgtcgt gccagctgca ttaacatggt catagctgtt tccttgcgta
     2461 ttgggcgctc tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc gggtaaagcc
     2521 tggggtgcct aatgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     2581 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     2641 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     2701 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     2761 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
     2821 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     2881 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     2941 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     3001 tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag
     3061 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     3121 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     3181 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     3241 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
     3301 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
     3361 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
     3421 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
     3481 ataccgcgag aaccacgctc accggctcca gatttatcag caataaacca gccagccgga
     3541 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
     3601 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
     3661 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
     3721 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
     3781 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
     3841 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
     3901 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
     3961 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
     4021 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
     4081 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
     4141 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
     4201 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
     4261 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt
     4321 ccccgaaaag tgccacctaa attgtaagcg ttaatatttt gttaaaattc gcgttaaatt
     4381 tttgttaaat cagctcattt tttaaccaat aggccgaaat cggcaaaatc ccttataaat
     4441 caaaagaata gaccgagata gggttgagtg gccgctacag ggcgctccca ttcgccattc
     4501 aggctgcgca actgttggga agggcgtttc ggtgcgggcc tcttcgctat tacgccagct
     4561 ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt tttcccagtc
     4621 acgacgttgt aaaacgacgg ccagtgagcg cgacgtaata cgactcacta tagggcgaat
     4681 tggcggaagg ccgtcaaggc cgtgacgtta aaactattaa gccatccaat cgaccgttag
     4741 tcgaatcagg accgctggtg cgagaagccg cgaagtatgg cgaatgcatc gtataacgtg
     4801 tggagtccgc tcattagagc gtcatgttta gacaagaaag ctacatattt aattgatccc
     4861 gatgatttta ttgataaatt gaccctaact ccatacacgg tattctacaa tggcggggtt
     4921 ttggtcaaaa tttccggact gcgattgtac atgctgttaa cggctccgcc cactattaat
     4981 gaaattaaaa attccaattt taaaaaacgc agcaagagaa acatttgtat gaaagaatgc
     5041 gtagaaggaa agaaaaatgt cgtggacatg ctgaacaaca agattaatat gcctccgtgt
     5101 ataaaaaaaa tattgaacga tttgaaagaa aacaatgtac cgcgcggcgg tatgtacagg
     5161 aagaggttta tactaaactg ttacattgca aacgtggttt cgtgtgccaa gtgtgaaaac
     5221 cgatgtttaa tcaaggctct gacgcatttc tacaaccacg actccaagtg tgtgggtgaa
     5281 gtcatgcatc ttttaatcaa atcccaagat gtgtataaac caccaaactg ccaaaaaatg
     5341 aaaactgtgg acaagctctg tccgtttgct ggcaactgca agggtctcaa tcctatttgt
     5401 aattattgaa taataaaaca attataaatg ctaaatttgt tttttattaa cgatacaaac
     5461 caaacgcaac aagaacattt gtagtattat ctataattga aaacgcgtag ttataatcgc
     5521 tgaggtaata tttaaaatca ttttcaaatg attcacagtt aatttgcgac aatataattt
     5581 tattttcaca taaactagac gccttgtcgt cttcttcttc gtattccttc tctttttcat
     5641 ttttctcctc ataaaaatta acatagttat tatcgtatcc atatatgtat ctatcgtata
     5701 gagtaaattt tttgttgtca taaatatata tgtctttttt aatggggtgt atagtaccgc
     5761 tgcgcatagt ttttctgtaa tttacaacag tgctattttc tggtagttct tcggagtgtg
     5821 ttgctttaat tattaaattt atataatcaa tgaatttggg atcgtcggtt ttgtacaata
     5881 tgttgccggc atagtacgca gcttcttcta gttcaattac accatttttt agcagcaccg
     5941 gattaacata actttccaaa atgttgtacg aaccgttaaa caaaaacagt tcacctccct
     6001 tttctatact attgtctgcg agcagttgtt tgttgttaaa aataacagcc attgtaatga
     6061 gacgcacaaa ctaatatcac aagctgccac cattttgttc gcccaggact ctagctatag
     6121 ttctagtggt tggctacgta tactccggaa tattaatag
//
